Homologous 28-kilodaltion immunodominant protein genes of Ehrlichia canis and uses thereof

Abstract

The present invention is directed to the cloning, sequencing and expression of homologous immunoreactive 28-kDa protein genes, p28-1, -2, -3, -5, -6, -7, -9, from a polymorphic multiple gene family of Ehrlichia canis. Further disclosed is a multigene locus encoding all nine homologous 28-kDa protein genes of Ehrlichia canis. Recombinant Ehrlichia canis 28-kDa proteins react with convalescent phase antiserum from an E. canis -infected dog, and may be useful in the development of vaccines and serodiagnostics that are particularly effective for disease prevention and serodiagnosis.

What is claimed is:  
     
         1 . An isolated DNA sequence encoding a 30-kilodalton protein of  Ehrlichia canis,  wherein said protein is immunoreactive with anti- Ehrlichia canis  serum.  
     
     
         2 . The DNA sequence of  claim 1 , wherein said protein comprises of an amino acid sequence selected from the group consisting of SEQ ID No. 2, 4, 6, 40, 42, 44 and 46.  
     
     
         3 . The DNA sequence of  claim 2 , wherein said protein has an N-terminal signal sequence.  
     
     
         4 . The DNA sequence of  claim 3 , wherein said protein is post-translationally modified to a 28-kilodalton protein.  
     
     
         5 . The DNA sequence of  claim 1 , wherein said DNA comprises a sequence selected from the group consisting of SEQ ID No. 1, 3, 5, 39, 41, 43 and 45.  
     
     
         6 . The DNA sequence of  claim 1 , wherein said DNA is contained in a single locus of  Ehrlichia canis.    
     
     
         7 . The DNA sequence of  claim 6 , wherein said locus is a multigene locus of 10,677 base pairs in length.  
     
     
         8 . The DNA sequence of  claim 7 , wherein said locus contains genes encoding homologous 28-kilodalton proteins of  Ehrlichia canis.    
     
     
         9 . The DNA sequence of  claim 8 , wherein said homologous 28-kilodalton proteins of  Ehrlichia canis  are selected from the group consisting of p28-1, p28-2, p 28 -3, p28-4, p28-5, p28-6, p28-7, p28-8 and p28-9.  
     
     
         10 . A vector comprising the DNA sequence of  claim 1 .  
     
     
         11 . The vector of  claim 10 , wherein said vector is an expression vector capable of expressing a peptide or polypeptide encoded by a sequence selected from the group consisting of SEQ ID No. 1, 3, 5, 39, 41, 43 and 45 when said expression vector is introduced into a cell.  
     
     
         12 . A recombinant protein comprises of an amino acid sequence selected from the group consisting of SEQ ID No. 2, 4, 6, 40, 42, 44 and 46.  
     
     
         13 . The recombinant protein of  claim 12 , wherein said amino acid sequence is encoded by a nucleic acid segment comprising a sequence selected from the group consisting of SEQ ID No. 1, 3, 5, 39, 41, 43 and 45.  
     
     
         14 . A host cell comprising a nucleic acid segment selected from the group consisting of SEQ ID No. 1, 3, 5, 39, 41, 43 and 45.  
     
     
         15 . A method of producing the recombinant protein of  claim 12 , comprising the steps of: 
 obtaining a vector that comprises an expression region comprising a sequence encoding the amino acid sequence selected from the group consisting of SEQ ID No. 2, 4, 6, 40, 42, 44 and 46 operatively linked to a promoter;    transfecting said vector into a cell; and    culturing said cell under conditions effective for expression of said expression region.    
     
     
         16 . An antibody immunoreactive with a polypeptide comprises of an amino acid sequence selected from the group consisting of SEQ ID No. 2, 4, 6, 40, 42, 44 and 46.  
     
     
         17 . A method of inhibiting  Ehrlichia canis  infection in a subject comprising the steps of: 
 identifying a subject prior to exposure or suspected of being exposed to or infected with  Ehrlichia canis;  and    administering a composition comprising a 28-kDa antigen of  Ehrlichia canis  in an amount effective to inhibit  Ehrlichia canis  infection.    
     
     
         18 . The method of  claim 17 , wherein said 28-kDa antigen is a recombinant protein comprising an amino acid sequence selected from the group consisting of SEQ ID No. 2, 4, 6, 40, 42, 44 and 46.  
     
     
         19 . The method of  claim 18 , wherein said recombinant protein is encoded by a gene comprising a sequence selected from the group consisting of SEQ ID No. 1, 3, 5, 39, 41, 43 and 45.  
     
     
         20 . The method of  claim 17 , wherein said composition comprising a 28-kDa antigen is dispersed in a pharmaceutically acceptable carrier.